Author response: Cryo-EM reveals distinct conformations of E. coli ATP synthase on exposure to ATP

ABSTRACTThe microcin PDI inhibits a diverse group of pathogenicEscherichia colistrains. Coculture of a single-gene knockout library (BW25113;n= 3,985 mutants) against a microcin PDI-producing strain (E. coli25) identified six mutants that were not susceptible (ΔatpA, ΔatpF, ΔdsbA, ΔdsbB, ΔompF, and ΔompR). Complementation of these genes restored susceptibility in all cases, and the loss of susceptibility was confirmed through independent gene knockouts inE. coliO157:H7 Sakai. Heterologous expression ofE. coliompFconferred susceptibility toSalmonella entericaandYersinia enterocoliticastrains that are normally unaffected by microcin PDI. The expression of chimeric OmpF and site-directed mutagenesis revealed that the K47G48N49region within the first extracellular loop ofE. coliOmpF is a putative binding site for microcin PDI. OmpR is a transcriptional regulator forompF, and consequently loss of susceptibility by the ΔompRstrain most likely is related to this function. Deletion of AtpA and AtpF, as well as AtpE and AtpH (missed in the original library screen), resulted in the loss of susceptibility to microcin PDI and the loss of ATP synthase function. Coculture of a susceptible strain in the presence of an ATP synthase inhibitor resulted in a loss of susceptibility, confirming that a functional ATP synthase complex is required for microcin PDI activity. Intransexpression ofompFin the ΔdsbAand ΔdsbBstrains did not restore a susceptible phenotype, indicating that these proteins are probably involved with the formation of disulfide bonds for OmpF or microcin PDI.

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Author response for "A rational approach to improving titer in E. coli ‐based cell‐free protein synthesis reactions"

10.1002/btpr.3062/v2/response1 ◽

2020 ◽

Author(s):

Noelle Colant ◽

Beatrice Melinek ◽

Jaime Teneb ◽

Stephen Goldrick ◽

William Rosenberg ◽

...

Keyword(s):

Protein Synthesis ◽

Author Response ◽

Rational Approach ◽

Free Protein ◽

E Coli ◽

Synthesis Reactions ◽

Cell Free Protein Synthesis

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The role of the amino‐terminal periplasmic domain of subunit a of the E. coli ATP synthase in function and assembly

The FASEB Journal ◽

10.1096/fasebj.20.4.a518-d ◽

2006 ◽

Vol 20 (4) ◽

Author(s):

Steven B Vik ◽

Robert R Ishmukhametov

Keyword(s):

Atp Synthase ◽

Subunit A ◽

E Coli ◽

Amino Terminal ◽

Periplasmic Domain

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Distances between the b-subunits in the tether domain of F0F1-ATP synthase from E. coli

Biochimica et Biophysica Acta (BBA) - Bioenergetics ◽

10.1016/j.bbabio.2005.03.013 ◽

2005 ◽

Vol 1708 (2) ◽

pp. 143-153 ◽

Cited By ~ 32

Author(s):

Stefan Steigmiller ◽

Michael Börsch ◽

Peter Gräber ◽

Martina Huber

Keyword(s):

Atp Synthase ◽

E Coli ◽

F0f1 Atp Synthase ◽

B Subunits

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Protein Sequence and Structure of N-terminal Amino Acids of Subunit Delta of Spinach Photosynthetic ATP-Synthase CF1

Zeitschrift für Naturforschung C ◽

10.1515/znc-1987-11-1215 ◽

1987 ◽

Vol 42 (11-12) ◽

pp. 1231-1238 ◽

Cited By ~ 11

Author(s):

Richard J. Berzborn ◽

Werner Finke ◽

Joachim Otto ◽

Helmut E . Meyer

Keyword(s):

Secondary Structure ◽

Atp Synthase ◽

Protein Sequence ◽

Alpha Helix ◽

Amino Acid Residues ◽

E Coli ◽

Helical Wheel ◽

Terminal Amino ◽

Amphipathic Alpha Helix ◽

Structure Calculations

Chloroplast ATP-synthase (CF1) subunit delta (δ) has been isolated from spinach thylakoids in the presence of SDS. By automated Edman degradation and online analysis of PTH derivatives the 35 N-terminal amino acid residues were sequenced. The mature protein starts with: NH2-Val-Asp-Ser-Thr-Ala-Ser-Arg-Tyr-Ala-. This protein sequence allows alignment of spinach δ with the sequences of Z. mays 25 kDa polypeptide, the δ subunit of Rps. blastica, Rsp. rubrum and E. coli F1, and of bovine OSCP, but not with mitochondrial δ. Secondary structure calculations and helical wheel plots reveal a conserved secondary structure. The analyzed N-terminal sequences probably build a short amphipathic alpha helix with two adjacent turns. The such aligned polar residues around Tyr8 of subunit δ are suitable to channel protons.

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The structure of the catalytic domain of the ATP synthase fromMycobacterium smegmatisis a target for developing antitubercular drugs

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1817615116 ◽

2019 ◽

Vol 116 (10) ◽

pp. 4206-4211 ◽

Cited By ~ 19

Author(s):

Alice Tianbu Zhang ◽

Martin G. Montgomery ◽

Andrew G. W. Leslie ◽

Gregory M. Cook ◽

John E. Walker

Keyword(s):

Atp Synthase ◽

Mycobacterium Smegmatis ◽

Catalytic Domain ◽

Atp Hydrolysis ◽

Atp Synthesis ◽

Geobacillus Stearothermophilus ◽

Antitubercular Drugs ◽

Atp Binding Site ◽

E Coli ◽

Catalytic Cycles

The crystal structure of the F1-catalytic domain of the adenosine triphosphate (ATP) synthase has been determined fromMycobacterium smegmatiswhich hydrolyzes ATP very poorly. The structure of the α3β3-component of the catalytic domain is similar to those in active F1-ATPases inEscherichia coliandGeobacillus stearothermophilus. However, its ε-subunit differs from those in these two active bacterial F1-ATPases as an ATP molecule is not bound to the two α-helices forming its C-terminal domain, probably because they are shorter than those in active enzymes and they lack an amino acid that contributes to the ATP binding site in active enzymes. InE. coliandG. stearothermophilus, the α-helices adopt an “up” state where the α-helices enter the α3β3-domain and prevent the rotor from turning. The mycobacterial F1-ATPase is most similar to the F1-ATPase fromCaldalkalibacillus thermarum, which also hydrolyzes ATP poorly. The βE-subunits in both enzymes are in the usual “open” conformation but appear to be occupied uniquely by the combination of an adenosine 5′-diphosphate molecule with no magnesium ion plus phosphate. This occupation is consistent with the finding that their rotors have been arrested at the same point in their rotary catalytic cycles. These bound hydrolytic products are probably the basis of the inhibition of ATP hydrolysis. It can be envisaged that specific as yet unidentified small molecules might bind to the F1domain inMycobacterium tuberculosis, prevent ATP synthesis, and inhibit the growth of the pathogen.

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Subunit II (b') and Not Subunit I (b) of Photosynthetic ATP Synthases is Equivalent to Subunit b of the ATP Synthases from Nonphotosynthetic Eubacteria. Evidence for a New Assignment of b-Type F0 Subunits

Zeitschrift für Naturforschung C ◽

10.1515/znc-1997-11-1211 ◽

1997 ◽

Vol 52 (11-12) ◽

pp. 789-798 ◽

Cited By ~ 1

Author(s):

Hans-Jürgen Tiburzy ◽

Richard J. Berzborn

Keyword(s):

Escherichia Coli ◽

Amino Acid ◽

Structural Feature ◽

Atp Synthase ◽

Primary Structure ◽

Structural Features ◽

Amino Acid Sequences ◽

E Coli ◽

Subunit B ◽

Atp Synthases

Abstract Subunit I of chloroplast ATP synthase is reviewed until now to be equivalent to subunit b of Escherichia coli ATP synthase, whereas subunit II is suggested to be an additional subunit in photosynthetic ATP synthases lacking a counterpart in E. coli. After publication of some sequences of subunits II a revision of this assignment is necessary. Based on the analysis of 51 amino acid sequences of b-type subunits concerning similarities in primary structure, isoelectric point and a discovered discontinuous structural feature, our data provide evidence that chloroplast subunit II (subunit b' of photosynthetic eubacteria) and not chloroplast subunit I (subunit b of photosynthetic eubacteria) is the equivalent of subunit b of nonphoto synthetic eubacteria, and therefore does have a counterpart in e.g. E. coli. In consequence, structural features essential for function should be looked for on subunit II (b').

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Western Faculty Profile: Dr. Derek McLachlin

Western Undergraduate Research Journal Health and Natural Sciences ◽

10.5206/wurjhns.2012-13.6 ◽

2012 ◽

Vol 3 (1) ◽

Author(s):

Ellen Y Xu ◽

Cheryl Y Yip

Keyword(s):

New York ◽

New York City ◽

Atp Synthase ◽

York City ◽

Assistant Professor ◽

Complex Mixtures ◽

Doctoral Studies ◽

E Coli ◽

Phosphorylated Peptides

Dr. Derek McLachlin is an assistant professor in the Department of Biochemistry at Western University. He obtained his PhD at Western University for his work on the quatenary structure of E. coli ATP synthase. Dr. McLachlin did his post-doctoral studies at Rockefeller University in New York City, where he analyzed phosphorylated peptides from complex mixtures. Ellen Xu and Cheryl Yip had the chance to interview him to learn more about his research and his time as a professor at Western.

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